Cell-free and cell-associated HIV-1 have been detected in semen from HIV-1-infected subjects. HIV-1 populations in semen have been shown to be different than those in blood of most infected subjects. We have recently demonstrated that the HIV-1 population in semen is further subcompartmentalized between seminal leukocytes and seminal plasma. Furthermore, HIV-1 in these two seminal subcompartments may originate from different anatomical sites of the male genital tract. However, there is no information available whether transmission occurs via cell-free or cell-associated HIV-1 or both present in semen. Compartmentalization of HIV-1 between seminal cells and seminal plasma raises the possibility that the viral dynamics in these two seminal compartments are different. Therefore, the responses of cell-free and cell- associated HIV-1 in semen to antiretroviral therapies could be different. Our overall hypothesis is that HIV- 1 in male genital organ is highly compartmentalized and this Compartmentalization can affect transmission and is sensitivity to therapy. To address these hypotheses we propose the following specific aims: 1) To determine whether cell-free or cell-associated HIV-1 in semen is involved in heterosexual transmission by analyzing the HIV-1 envelope gene from seminal cells and seminal plasma from transmitting men with and without therapy, and blood from their infected female partners soon after exposure;2) To investigate on the effect of short-term and long-term therapy on cell-associated and cell-free HIV-1 in semen. Clearance rate of cell-free and cell-associated HIV-1 in semen and blood will be measured during the short-term therapy by measuring viral load at short intervals before and after therapy. Long term therapy (more than 2 years) will be used to determine whether antiretroviral therapy has a differential effect on the viral reservoir in cell-free compartment as compared to cell-associated compartment in semen and blood. This will be done by measuring viral expression in cultured cells and HIV-1 evolution;3) To determine the anatomical location of HIV-1 in the male genital organ and evaluate the effect of antiretroviral therapy on HIV- 1 in the various male genital tissues. The level of viral RNA in prostate, seminal vesicle, testes, vas-deference and urethra in HIV-1-infected subjects before and after therapy will be measured. We have assembled all the resources and expertise to accomplish the stated specific aims in this proposal. Semen and blood sample for studies described in Aims 1 and 2 will be obtained from HIV infected subjects from Dominican republic and Puerto Rico though our ongoing collaborative studies. Tissue sample for Aim 3 will be obtained from the National Disease Research Interchange (NDRI).The information from the proposed study will be extremely useful to our understanding of transmission and eradication of HIV-1 following therapy.